Engineered Microencapsulated Lactoferrin Nanoconjugates for Oral Targeted Treatment of Colon Cancer.

Despite current progress in the development of targeted therapies for cancer treatment, there is a lack in convenient therapeutics for colorectal cancer (CRC). Lactoferrin nanoparticles (Lf NPs) are a promising drug delivery system in cancer therapy. However, numerous obstacles impede their oral delivery, including instability against stomach enzymes and premature uptake during passage through the small intestine. Microencapsulation of Lf NPs offer a great solution for these obstacles. It can protect Lf NPs and their drug payloads from degradation in the upper gastrointestinal tract (GIT), reduce burst drug release, and improve the release profile of the encapsulated NPs triggered by stimuli in the colon. Here, we developed nanoparticle-in-microparticle delivery systems (NIMDs) for the oral delivery of docetaxel (DTX) and atorvastatin (ATR). The NPs were obtained by dual conjugation of DTX and ATR into the Lf backbone, which was further microencapsulated into calcium-crosslinked microparticles using polysaccharide-protein hybrid copolymers. The NIMDs showed no detectable drug release in the upper GIT compared to NPs. Furthermore, sustained release of the NPs from the NIMDs in rat cecal content was observed. Moreover, the in vivo study demonstrated the superiority of the NIMDs over NPs in CRC treatment by suppressing p-AKT, p-ERK1/2, and NF-κB. This study provides the proof of concept for using NIMDs to enhance the effect of protein NPs in CRC treatment.

Promoted Antitumor Activity of Myricetin against Lung Carcinoma Via Nanoencapsulated Phospholipid Complex in Respirable Microparticles.

PURPOSE: Myricetin (MYR) flavonoid is well-recognized for its antioxidant, anti-inflammatory and anti-tumor potential. Introducing nanomedicine was the ultimate resort to solve the imperfections of this nutraceutical, namely solubility, stability and delivery issues. The study, thus, aims at developing inhalable microparticles comprising MYR solid lipid nanoparticles (SLNs) for lung cancer therapy. METHODS: A two-step preparation procedure starting with complexation of MYR with the phospholipid Lipoid-S100, followed by nanoencapsulation in Gelucire-based, surfactant-free SLNs was developed. SLNs were characterized in terms of physicochemical properties, MYR loading, release behavior as well as anti-tumor potential and cellular uptake. Respirable microparticles were then obtained by spray drying SLNs with carbohydrate carriers. Their size, flowability and pulmonary deposition pattern were assessed. RESULTS: Optimized SLNs were 75.98 nm in diameter with a zeta-potential of -22.5 mV, and an encapsulation efficiency of 84.5%. Attempts to ameliorate drug loading implicate MYR-phospholipid complexation (MYR-PH-CPX) prior to its entrapment in SLNs, which ensured 5-fold increase in drug loading. Viability assays were modified to guarantee MYR chemical stability. Superior antitumor activity of MYR-phospholipid-complex and 3-fold reduction in IC50 were accomplished with MYR-SLNs. This could be related to enhanced cellular uptake revealed by confocal imaging and doubled fluorescence intensity. SLNs entrapping MYR-PH-CPX were spray-dried with carbohydrate carriers to produce respirable microparticles. The latter ensured MMAD of 2.39 µm and span index of 1.84, in addition to good flowability and > 80% release over 8 h. Deposition experiments revealed MMAD of 2.77 µm, FPF of 81.23 and EF of 93% indicating particle deposition in the targeted bronchial region. CONCLUSIONS: The study highlights the ability of phospholipid-complex on the nanoencapsulation, cellular uptake and antitumor activity of MYR. Formulation of respirable microparticles gives promises of efficacious therapy of lung carcinoma.

Oleic acid-based nanosystems for mitigating acute respiratory distress syndrome in mice through neutrophil suppression: how the particulate size affects therapeutic efficiency.

BACKGROUND: Oleic acid (OA) is reported to show anti-inflammatory activity toward activated neutrophils. It is also an important material in nanoparticles for increased stability and cellular internalization. We aimed to evaluate the anti-inflammatory activity of injectable OA-based nanoparticles for treating lung injury. Different sizes of nanocarriers were prepared to explore the effect of nanoparticulate size on inflammation inhibition. RESULTS: The nanoparticles were fabricated with the mean diameters of 105, 153, and 225 nm. The nanocarriers were ingested by isolated human neutrophils during a 5-min period, with the smaller sizes exhibiting greater uptake. The size reduction led to the decrease of cell viability and the intracellular calcium level. The OA-loaded nanosystems dose-dependently suppressed the superoxide anion and elastase produced by the stimulated neutrophils. The inhibition level was comparable for the nanoparticles of different sizes. In the ex vivo biodistribution study, the pulmonary accumulation of nanoparticles increased following the increase of particle size. The nanocarriers were mainly excreted by the liver and bile clearance. Mice were exposed to intratracheal lipopolysaccharide (LPS) to induce acute respiratory distress syndrome (ARDS), like lung damage. The lipid-based nanocarriers mitigated myeloperoxidase (MPO) and cytokines more effectively as compared to OA solution. The larger nanoparticles displayed greater reduction on MPO, TNF-α, and IL-6 than the smaller ones. The histology confirmed the decreased pulmonary neutrophil recruitment and lung-architecture damage after intravenous administration of larger nanoparticles. CONCLUSIONS: Nanoparticulate size, an essential property governing the anti-inflammatory effect and lung-injury therapy, had different effects on activated neutrophil inhibition and in vivo therapeutic efficacy.

Combining hydrophilic chemotherapy and hydrophobic phytotherapy via tumor-targeted albumin-QDs nano-hybrids: covalent coupling and phospholipid complexation approaches.

BACKGROUND: The rationale of this study is to combine the merits of both albumin nanoparticles and quantum dots (QDs) in improved drug tumor accumulation and strong fluorescence imaging capability into one carrier. However, premature drug release from protein nanoparticles and high toxicity of QDs due to heavy metal leakage are among challenging hurdles. Following this platform, we developed cancer nano-theranostics by coupling biocompatible albumin backbone to CdTe QDs and mannose moieties to enhance tumor targeting and reduce QDs toxicity. The chemotherapeutic water soluble drug pemetrexed (PMT) was conjugated via tumor-cleavable bond to the albumin backbone for tumor site-specific release. In combination, the herbal hydrophobic drug resveratrol (RSV) was preformulated as phospholipid complex which enabled its physical encapsulation into albumin nanoparticles. RESULTS: Albumin-QDs theranostics showed enhanced cytotoxicity and internalization into breast cancer cells that could be traced by virtue of their high fluorescence quantum yield and excellent imaging capacity. In vivo, the nanocarriers demonstrated superior anti-tumor effects including reduced tumor volume, increased apoptosis, and inhibited angiogenesis in addition to non-immunogenic response. Moreover, in vivo bioimaging test demonstrated excellent tumor-specific accumulation of targeted nanocarriers via QDs-mediated fluorescence. CONCLUSION: Mannose-grafted strategy and QD-fluorescence capability were beneficial to deliver albumin nanocarriers to tumor tissues and then to release the anticancer drugs for killing cancer cells as well as enabling tumor imaging facility. Overall, we believe albumin-QDs nanoplatform could be a potential nano-theranostic for bioimaging and targeted breast cancer therapy.

Lactobionic/Folate Dual-Targeted Amphiphilic Maltodextrin-Based Micelles for Targeted Codelivery of Sulfasalazine and Resveratrol to Hepatocellular Carcinoma.

In this study, promising approaches of dual-targeted micelles and drug-polymer conjugation were combined to enable injection of poorly soluble anticancer drugs together with site-specific drug release. Ursodeoxycholic acid (UDCA) as a hepatoprotective agent was grafted to maltodextrin (MD) via carbodiimide coupling to develop amphiphilic maltodextrin-ursodeoxycholic acid (MDCA)-based micelles. Sulfasalazine (SSZ), as a novel anticancer agent, was conjugated via a tumor-cleavable ester bond to MD backbone to obtain tumor-specific release, whereas resveratrol (RSV) was physically entrapped within the hydrophobic micellar core. For maximal tumor-targeting, both folic acid (FA) and lactobionic acid (LA) were coupled to the surface of micelles to obtain dual-targeted micelles. The decrease of critical micelle concentration (CMC) from 0.012 to 0.006 mg/mL declares the significance of a dual hydrophobicized core of micelles by both UDCA and SSZ. The dual-targeted micelles showed a great hemocompatibility, as well as enhanced cytotoxicity and internalization into HepG-2 liver cancer cells via binding to overexpressed folate and asialoglycoprotein receptors. In vivo, the micelles demonstrated superior antitumor effects revealed as reduction in the liver/body weight ratio, inhibition of angiogenesis, and enhanced apoptosis. Overall, combined strategies of dual active targeted micelles with bioresponsive drug conjugation could be utilized as a promising approach for tumor-targeted drug delivery.

Use of cilomilast-loaded phosphatiosomes to suppress neutrophilic inflammation for attenuating acute lung injury: the effect of nanovesicular surface charge.

BACKGROUND: Cilomilast is a phosphodiesterase 4 (PDE4) inhibitor for treating inflammatory lung diseases. This agent has a narrow therapeutic index with significant adverse effects on the nervous system. This study was conducted to entrap cilomilast into PEGylated phosphatidylcholine-rich niosomes (phosphatiosomes) to improve pulmonary delivery via the strong affinity to pulmonary surfactant film. Neutrophils were used as a cell model to test the anti-inflammatory activity of phosphatiosomes. In an in vivo approach, mice were given lipopolysaccharide to produce acute lung injury. The surface charge in phosphatiosomes that influenced the anti-inflammatory potency is discussed in this study. RESULTS: The average diameter of the phosphatiosomes was about 100 nm. The zeta potential of anionic and cationic nanovesicles was - 35 and 32 mV, respectively. Cilomilast in both its free and nanocapsulated forms inhibited superoxide anion production but not elastase release in activated neutrophils. Cationic phosphatiosomes mitigated calcium mobilization far more effectively than the free drug. In vivo biodistribution evaluated by organ imaging demonstrated a 2-fold ameliorated lung uptake after dye encapsulation into the phosphatiosomes. The lung/brain distribution ratio increased from 3 to 11 after nanocarrier loading. The intravenous nanocarriers deactivated the neutrophils in ALI, resulting in the elimination of hemorrhage and alveolar wall damage. Only cationic phosphatiosomes could significantly suppress IL-1ß and TNF-α in the inflamed lung tissue. CONCLUSIONS: These results suggest that phosphatiosomes should further be investigated as a potential nanocarrier for the treatment of pulmonary inflammation.

Flutamide-Loaded Zein Nanocapsule Hydrogel, a Promising Dermal Delivery System for Pilosebaceous Unit Disorders.

Zein is a naturally occurring corn protein having similarity to skin keratin. Owing to its hydrophobicity and biodegradability, zein nanocarriers are promising drug delivery vehicles for hydrophobic dermatological drugs. In this study, zein-based nanocapsules (ZNCs) were exploited for the first time as dermal delivery carriers for flutamide (FLT), an antiandrogen used for the management of pilosebasceous unit disorders. FLT-loaded ZNC of appropriate particle size and negative surface charge were prepared by nanoprecipitation method. The dermal permeation and skin retention of FLT from ZNCs were studied in comparison to corresponding nanoemulsion (NE) and hydroalcoholic drug solution (HA). ZNCs showed a significantly lower permeation flux compared to NE and HA while increasing the skin retention of FLT. Confocal laser scanning microscopy (CLSM) demonstrated the follicular localization of the fluorescently labeled NCs. The incorporation of NCs in chitosan gel or Carbomer® 934 gel was studied. Carbomer® gel increased the skin retention of FLT compared to chitosan gel. Accordingly, Carbomer® hydrogel embedding FLT-loaded ZNCs is a promising inexpensive, biocompatible dermal delivery nanocarrier for localized therapy of PSU disorders suitable for application on oily skin.

Multi-Reservoir Phospholipid Shell Encapsulating Protamine Nanocapsules for Co-Delivery of Letrozole and Celecoxib in Breast Cancer Therapy.

PURPOSE: In the current work, we propose a combined delivery nanoplatform for letrozole (LTZ) and celecoxib (CXB). METHODS: Multi-reservoir nanocarriers were developed by enveloping protamine nanocapsules (PRM-NCs) within drug-phospholipid complex bilayer. RESULTS: Encapsulation of NCs within phospholipid bilayer was confirmed by both size increase from 109.7 to 179.8 nm and reduction of surface charge from +19.0 to +7.78 mV. The multi-compartmental core-shell structure enabled biphasic CXB release with initial fast release induced by complexation with phospholipid shell followed by prolonged release from oily core. Moreover, phospholipid coating provided protection for cationic PRM-NCs against interaction with RBCs and serum proteins enabling their systemic administration. Pharmacokinetic analysis demonstrated prolonged circulation and delayed clearance of both drugs after intravenous administration into rats. The superior anti-tumor efficacy of multi-reservoir NCs was manifested as powerful cytotoxicity against MCF-7 breast cancer cells and marked reduction in the mammary tumor volume in Ehrlich ascites bearing mice compared with free LTZ-CXB combination. Moreover, the NCs induced apoptotic caspase activation and marked inhibition of aromatase expression and angiogenic marker, VEGF as well as inhibition of both NFκB and TNFα. CONCLUSIONS: Multi-reservoir phospholipid shell coating PRM-NCs could serve as a promising nanocarrier for parenteral combined delivery of LTZ and CXB.

Recent advances in nano-based drug delivery systems for treatment of liver cancer.

Liver cancer is one of the aggressive primary tumors as evident by high rate of incidence and mortality. Conventional treatments (e.g. chemotherapy) suffer from various drawbacks including wide drug distribution, low localized drug concentration, and severe off-site toxicity. Therefore, they cannot satisfy the mounting need for safe and efficient cancer therapeutics, and alternative novel strategies are needed. Nano-based drug delivery systems (NDDSs) are among these novel approaches that can improve the overall therapeutic outcomes. NDDSs are designed to encapsulate drug molecules and target them specifically to liver cancer. Thus, NDDSs can selectively deliver therapeutic agents to the tumor cells and avoid distribution to off-target sites which should improve the safety profile of the active agents. Nonetheless, NDDSs should be well designed, in terms of the preparing materials, nanocarriers structure, and the targeting strategy, in order to accomplish these objectives. This review discusses the latest advances of NDDSs for cancer therapy with emphasis on the aforementioned essential design components. The review also entails the challenges associated with the clinical translation of NDDSs, and the future perspectives towards next-generation NDDSs.

Modulating tumoral exosomes and fibroblast phenotype using nanoliposomes augments cancer immunotherapy.

Cancer cells program fibroblasts into cancer associated fibroblasts (CAFs) in a two-step manner. First, cancer cells secrete exosomes to program quiescent fibroblasts into activated CAFs. Second, cancer cells maintain the CAF phenotype via activation of signal transduction pathways. We rationalized that inhibiting this two-step process can normalize CAFs into quiescent fibroblasts and augment the efficacy of immunotherapy. We show that cancer cell-targeted nanoliposomes that inhibit sequential steps of exosome biogenesis and release from lung cancer cells block the differentiation of lung fibroblasts into CAFs. In parallel, we demonstrate that CAF-targeted nanoliposomes that block two distinct nodes in fibroblast growth factor receptor (FGFR)-Wnt/ß-catenin signaling pathway can reverse activate CAFs into quiescent fibroblasts. Co-administration of both nanoliposomes significantly improves the infiltration of cytotoxic T cells and enhances the antitumor efficacy of αPD-L1 in immunocompetent lung cancer-bearing mice. Simultaneously blocking the tumoral exosome-mediated activation of fibroblasts and FGFR-Wnt/ß-catenin signaling constitutes a promising approach to augment immunotherapy.

Engineering nanomedicines for immunogenic eradication of cancer cells: Recent trends and synergistic approaches.

Resistance to cancer immunotherapy is mainly attributed to poor tumor immunogenicity as well as the immunosuppressive tumor microenvironment (TME) leading to failure of immune response. Numerous therapeutic strategies including chemotherapy, radiotherapy, photodynamic, photothermal, magnetic, chemodynamic, sonodynamic and oncolytic therapy, have been developed to induce immunogenic cell death (ICD) of cancer cells and thereby elicit immunogenicity and boost the antitumor immune response. However, many challenges hamper the clinical application of ICD inducers resulting in modest immunogenic response. Here, we outline the current state of using nanomedicines for boosting ICD of cancer cells. Moreover, synergistic approaches used in combination with ICD inducing nanomedicines for remodeling the TME via targeting immune checkpoints, phagocytosis, macrophage polarization, tumor hypoxia, autophagy and stromal modulation to enhance immunogenicity of dying cancer cells were analyzed. We further highlight the emerging trends of using nanomaterials for triggering amplified ICD-mediated antitumor immune responses. Endoplasmic reticulum localized ICD, focused ultrasound hyperthermia, cell membrane camouflaged nanomedicines, amplified reactive oxygen species (ROS) generation, metallo-immunotherapy, ion modulators and engineered bacteria are among the most innovative approaches. Various challenges, merits and demerits of ICD inducer nanomedicines were also discussed with shedding light on the future role of this technology in improving the outcomes of cancer immunotherapy.

Nanomedicines as enhancers of tumor immunogenicity to augment cancer immunotherapy.

The potential of cancer immunotherapy is hampered by the poor immunogenicity of cancer cells. Strategies to enhance tumor immunogenicity are imperative to enhance T cell-mediated anti-tumor immunity. Although conventional therapeutics can increase tumor antigen expression or stimulate the release of danger signals to promote immunogenic cell death (ICD), they face challenges relating to efficacy and tumor-specific delivery. Nanomedicines can efficiently deliver tumor antigens, immune adjuvants, epigenetic modulators, or ICD inducers through targeted drug delivery with minimal off-target effects. Collectively, nanomedicines can overcome biological barriers to immunotherapy through targeted antigen delivery, induction of ICD, or epigenetic remodeling, resulting in increased tumor immunogenicity.

Novel spray-dried genipin-crosslinked casein nanoparticles for prolonged release of alfuzosin hydrochloride.

PURPOSE: To propose a simple method for the development of genipin-crosslinked casein micelles as a new delivery platform for prolonged release of alfuzosin hydrochloride. METHODS: Crosslinked casein micelles entrapping alfuzosin were transformed into solid redispersible nanoparticles via spray-drying technique with no need for drying adjuvants based on the stabilizing effect of casein. RESULTS: The nanoparticles displayed high production yields (86.99-94.63% w/w) with a reasonable drug incorporation efficiency ranged from 92.86 to 97.75%. The nanoparticles were readily reconstituted in aqueous solution with a particle size range of 122.1-260.0 nm and a zeta potential range of -21.6 to -36.6 mV indicating a good colloidal stability. No drug crystals were detectable in the scanning electron micrographs revealing successful encapsulation of alfuzosin into casein nanoparticles which was confirmed by differential scanning calorimetry. The nanoparticles succeeded in prolonging the drug release that could be controlled by modulating the genipin crosslinking degree. The release data showed a good fit into Higuchi release kinetics with non-Fickian type of drug diffusion. CONCLUSIONS: These results demonstrated that genipin-crosslinking combined with spray-drying technique could be used as a promising tool to develop solid redispersible casein nanoparticles with sustained drug release properties.

Micellar delivery of flutamide via milk protein nanovehicles enhances its anti-tumor efficacy in androgen-dependent prostate cancer rat model.

PURPOSE: This article describes the preparation, physicochemical characterization and in vivo assessment of parenteral colloidal formulation of flutamide (FLT) based on biocompatible casein (CAS) self-assembled micelles in order to control drug release, enhance its antitumor efficacy and reduce its hepatotoxicity. METHODS: Spray-drying technique was successfully utilized to obtain solidified redispersible drug-loaded micelles. RESULTS: Spherical core-shell micelles were obtained with a particle size below 100 nm and a negative zeta potential above -30 mV exhibiting a sustained drug release up to 5 days. After intravenous administration into prostate cancer bearing rats for 28 days, FLT-loaded CAS micelles showed a higher antitumor efficacy as revealed by significantly higher reduction in PSA serum level (65.95%) compared to free FLT (55.43%). Moreover, micellar FLT demonstrated a marked decrease in relative weights of both prostate tumor and seminal vesicle (34.62 and 24.59%) compared to free FLT (11.86 and 17.74%), respectively. These antitumor responses were associated with notable reduction of cell proliferation, intratumoral angiogenesis and marked increase of tumor apoptosis. A significantly lower risk of hepatotoxicity was observed by micellar FLT as evidenced by lower alanine aminotransferase (ALT) serum level compared to free FLT. CONCLUSIONS: Overall this approach suggested that CAS micelles might be an ideal candidate for intravenous delivery of hydrophobic anticancer drugs.

Engineered nanomedicines to overcome resistance of pancreatic cancer to immunotherapy.

Pancreatic cancer (PC) is a highly aggressive malignant type of cancer. Although immunotherapy has been successfully used for treatment of many cancer types, many challenges limit its success in PC. Therefore, nanomedicines were engineered to enhance the responsiveness of PC cells to immune checkpoint inhibitors (ICIs). In this review, we highlight recent advances in engineering nanomedicines to overcome PC immune resistance. Nanomedicines were used to increase the immunogenicity of PC cells, inactivate stromal cancer-associated fibroblasts (CAFs), enhance the antigen-presenting capacity of dendritic cells (DCs), reverse the highly immunosuppressive nature of the tumor microenvironment (TME), and, hence, improve the infiltration of cytotoxic T lymphocytes (CTLs), resulting in efficient antitumor immune responses.

Engineered Sericin-Tagged Layered Double Hydroxides for Combined Delivery of Pemetrexed and ZnO Quantum Dots as Biocompatible Cancer Nanotheranostics.

AIM: Despite extensive progress in the field of cancer nanotheranostics, clinical development of biocompatible theranostic nanomedicine remains a formidable challenge. Herein, we engineered biocompatible silk-sericin-tagged inorganic nanohybrids for efficient treatment and imaging of cancer cells. The developed nanocarriers are anticipated to overcome the premature release of the chemotherapeutic drug pemetrexed (PMX), enhance the colloidal stability of layered double hydroxides (LDHs), and maintain the luminescence properties of ZnO quantum dots (QDs). Materials and Methods: PMX-intercalated LDHs were modified with sericin and coupled to ZnO QDs for therapy and imaging of breast cancer cells. Results: The optimized nanomedicine demonstrated a sustained release profile of PMX, and high cytotoxicity against MDA-MB-231 cells compared to free PMX. In addition, high cellular uptake of the engineered nanocarriers into MDA-MB-231 breast cancer cells was accomplished. Conclusions: Conclusively, the LDH-sericin nanohybrids loaded with PMX and conjugated to ZnO QDs offered a promising cancer theranostic nanomedicine.

Combined Cancer Immunotheranostic Nanomedicines: Delivery Technologies and Therapeutic Outcomes.

Cancer is among the main causes of mortality all over the world. The delayed diagnosis is directly related to the decrease in survival rate. The use of immunotherapy has dramatically changed the treatment outcomes of different types of cancers. However, many patients still do not respond to immunotherapies, and many also suffer from severe immune-related side effects. Recent advances in the fields of nanomedicine bioengineering and in particular imaging offered new approaches which can enhance not only the safety but also the efficacy of immunotherapy. Theranostics has showed great progress as a branch of medicine which integrates both diagnosis and therapy in a single system. The outcomes from animal studies demonstrated an improvement in the diagnostic and immunotherapeutic potential of nanoparticles within the theranostic framework. Herein, we discuss the most recent developments in the application of nanotheranostics for combining tumor imaging and cancer immunotherapies.

Engineered nanomedicines for augmenting the efficacy of colorectal cancer immunotherapy.

Colorectal cancer (CRC) is one of the most devastating diseases worldwide. Immunotherapeutic agents for CRC treatment have shown limited efficacy due to the immunosuppressive tumor microenvironment (TME). In this context, various types of nanoparticles (NPs) have been used to reverse the immunosuppressive TME, potentiate the effect of immunotherapeutic agents and reduce their systemic side effects. Many advantages could be offered by NPs, related to drug-loading efficiency, particle size and others that can potentially aid the delivery of immunotherapeutic agents. The recent research on how nano-based immunotherapy can remodel the immunosuppressive TME of CRC and hence boost the antitumor immune response, as well as the challenges that face clinical translation of NPs and future perspectives, are summarized in this review article.

Methotrexate-Lactoferrin Targeted Exemestane Cubosomes for Synergistic Breast Cancer Therapy.

While the treatment regimen of certain types of breast cancer involves a combination of hormonal therapy and chemotherapy, the outcomes are limited due to the difference in the pharmacokinetics of both treatment agents that hinders their simultaneous and selective delivery to the cancer cells. Herein, we report a hybrid carrier system for the simultaneous targeted delivery of aromatase inhibitor exemestane (EXE) and methotrexate (MTX). EXE was physically loaded within liquid crystalline nanoparticles (LCNPs), while MTX was chemically conjugated to lactoferrin (Lf) by carbodiimide reaction. The anionic EXE-loaded LCNPs were then coated by the cationic MTX-Lf conjugate via electrostatic interactions. The Lf-targeted dual drug-loaded LCNPs exhibited a particle size of 143.6 ± 3.24 nm with a polydispersity index of 0.180. It showed excellent drug loading with an EXE encapsulation efficiency of 95% and an MTX conjugation efficiency of 33.33%. EXE and MTX showed synergistic effect against the MCF-7 breast cancer cell line with a combination index (CI) of 0.342. Furthermore, the Lf-targeted dual drug-loaded LCNPs demonstrated superior synergistic cytotoxic activity with a combination index (CI) of 0.242 and a dose reduction index (DRI) of 34.14 and 4.7 for EXE and MTX, respectively. Cellular uptake studies demonstrated higher cellular uptake of Lf-targeted LCNPs into MCF-7 cancer cells than non-targeted LCNPs after 4 and 24 h. Collectively, the targeted dual drug-loaded LCNPs are a promising candidate offering combinational hormonal therapy/chemotherapy for breast cancer.

Sequential Delivery of Novel Triple Drug Combination via Crosslinked Alginate/Lactoferrin Nanohybrids for Enhanced Breast Cancer Treatment.

While breast cancer remains a global health concern, the elaboration of rationally designed drug combinations coupled with advanced biocompatible delivery systems offers new promising treatment venues. Herein, we repurposed rosuvastatin (RST) based on its selective tumor apoptotic effect and combined it with the antimetabolite pemetrexed (PMT) and the tumor-sensitizing polyphenol honokiol (HK). This synergistic three-drug combination was incorporated into protein polysaccharide nanohybrids fabricated by utilizing sodium alginate (ALG) and lactoferrin (LF), inspired by the stealth property of the former and the cancer cell targeting capability of the latter. ALG was conjugated to PMT and then coupled with LF which was conjugated to RST, forming core shell nanohybrids into which HK was physically loaded, followed by cross linking using genipin. The crosslinked HK-loaded PMT-ALG/LF-RST nanohybrids exhibited a fair drug loading of 7.86, 5.24 and 6.11% for RST, PMT and HK, respectively. It demonstrated an eight-fold decrease in the IC50 compared to the free drug combination, in addition to showing an enhanced cellular uptake by MCF-7 cells. The in vivo antitumor efficacy in a breast cancer-bearing mouse model confirmed the superiority of the triple cocktail-loaded nanohybrids. Conclusively, our rationally designed triple drug-loaded protein/polysaccharide nanohybrids offer a promising, biocompatible approach for an effective breast tumor suppression.